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Antibody Sampler Kit Heat Shock Protein Binding

The HSP/Chaperone Sampler Kit provides an economical means to investigate protein folding within the cell. The kit contains enough primary and secondary antibodies to perform two Western blot experiments with each antibody.
The Exosomal Marker Antibody Sampler Kit provides an economical means to evaluate the presence of exosomal markers. The kit includes enough primary antibody to perform two western blot experiments for each target.
The Pro-Apoptosis Bcl-2 Family Antibody Sampler Kit II provides an economical means to examine several members of the Bcl-2 family. The kit contains enough primary antibody to perform two western blot experiments.

Background: The Bcl-2 family consists of a number of evolutionarily conserved proteins containing Bcl-2 homology domains (BH) that regulate apoptosis through control of mitochondrial membrane permeability and release of cytochrome c (1-3). Four BH domains have been identified (BH1-4) that mediate protein interactions. The family can be separated into three groups based upon function and sequence homology: pro-survival members include Bcl-2, Bcl-xL, Mcl-1, A1 and Bcl-w; pro-apoptotic proteins include Bax, Bak and Bok; and "BH3 only" proteins Bad, Bik, Bid, Puma, Bim, Bmf, Noxa and Hrk. Interactions between death-promoting and death-suppressing Bcl-2 family members has led to a rheostat model in which the ratio of pro-apoptotic and anti-apoptotic proteins controls cell fate (4). Thus, pro-survival members exert their behavior by binding to and antagonizing death-promoting members. In general, the "BH3-only members" can bind to and antagonize the pro-survival proteins leading to increased apoptosis (5). While some redundancy of this system likely exists, tissue specificity, transcriptional and post-translational regulation of many of these family members can account for distinct physiological roles.

The Pro-Apoptosis Bcl-2 Family Antibody Sampler Kit provides an economical means to examine several members of the Bcl-2 family and their activation status. The kit contains enough primary and secondary antibodies to perform two Western blot experiments per primary antibody.

Background: The Bcl-2 family consists of a number of evolutionarily conserved proteins containing Bcl-2 homology domains (BH) that regulate apoptosis through control of mitochondrial membrane permeability and release of cytochrome c (1-3). Four BH domains have been identified (BH1-4) that mediate protein interactions. The family can be separated into three groups based upon function and sequence homology: pro-survival members include Bcl-2, Bcl-xL, Mcl-1, A1 and Bcl-w; pro-apoptotic proteins include Bax, Bak and Bok; and "BH3 only" proteins Bad, Bik, Bid, Puma, Bim, Bmf, Noxa and Hrk. Interactions between death-promoting and death-suppressing Bcl-2 family members has led to a rheostat model in which the ratio of pro-apoptotic and anti-apoptotic proteins controls cell fate (4). Thus, pro-survival members exert their behavior by binding to and antagonizing death-promoting members. In general, the "BH3-only members" can bind to and antagonize the pro-survival proteins leading to increased apoptosis (5). While some redundancy of this system likely exists, tissue specificity, transcriptional and post-translational regulation of many of these family members can account for distinct physiological roles.

The Huntingtin Interaction Antibody Sampler kit provides an economical means of detecting transcription-related proteins that interact with Huntingtin (Htt). This kit contains enough antibody to perform two western blot experiments per primary antibody.
The NF-κB Pathway Sampler Kit contains reagents to examine the activation state and total protein levels of key proteins in the NF-κB pathway: IKKα, IKKβ, NF-κB p65/RelA and IκBα. The kit contains enough primary and secondary antibodies to perform two Western blot experiments per primary antibody.

Background: The transcriptional nuclear factor κB (NF-κB)/Rel transcription factors are present in the cytosol in an inactive state, complexed with the inhibitory IκB proteins. Activation occurs via phosphorylation of IκBα at Ser32 and Ser36, resulting in the ubiquitin-mediated proteasome-dependent degradation of IκBα and the release and nuclear translocation of active NF-κB dimers. The regulation of IκBβ and IκBε is similar to that of IκBα, however, the phosphorylation and degradation of these proteins occurs with much slower kinetics. Phosphorylation of IκBβ occurs at Ser/Thr19 and Ser23, while IκBε can be phosphorylated at Ser18 and Ser22. The key regulatory step in this pathway involves activation of a high molecular weight IkappaB kinase (IKK) complex, consisting of three tightly associated IKK subunits. IKKα and IKKβ serve as the catalytic subunits of the kinase. Activation of IKK depends on phosphorylation at Ser177 and Ser181 in the activation loop of IKKβ (176 and 180 in IKKα). NF-κB-inducing kinase (NIK), TANK-binding kinase 1 (TBK1), and its homolog IKKε (IKKi), phosphorylate and activate IKKα and IKKβ.

The Tricarboxylic Acid Cycle Sampler Kit provides an economical means of detecting select components involved in tricarboxylic acid cycle. The kit contains enough primary antibodies to perform at least two western blot experiments per antibody.

Background: The tricarboxylic acid (TCA) cycle includes various enzymatic reactions that constitute a key part of cellular aerobic respiration. The transport of the glycolytic end product pyruvate into mitochondria and the decarboxylation of pyruvate in the TCA cycle generate energy through oxidative phosphorylation under aerobic conditions (1,2). Two inner mitochondrial membrane proteins, mitochondrial pyruvate carrier 1 (MPC1) and mitochondrial pyruvate carrier 2 (MPC2), form a 150 kDa complex and are essential proteins in the facilitated transport of pyruvate into mitochondria (1,2). Citrate synthase catalyzes the first and rate-limiting reaction of the TCA cycle (3). Mitochondrial aconitase 2 (ACO2) catalyzes the conversion of citrate to isocitrate via cis-aconitate (4). IDH1 and IDH2 are two of the three isocitrate dehydrogenases that catalyze oxidative decarboxylation of isocitrate to α-ketoglutarate (α-KG) (5). IDH1 functions as a tumor suppressor in the cytoplasm and peroxisomes, whereas IDH2 is in mitochondria and is involved in the TCA cycle (5). Mutations in IDH2 have also been identified in malignant gliomas (6). Dihydrolipoamide succinyltransferase (DLST) is a subunit of the α-ketoglutarate dehydrogenase complex, a key enzymatic complex in the TCA cycle (7). Succinate dehydrogenase subunit A (SDHA) is a component of the TCA cycle and the electron transport chain and is involved in the oxidation of succinate (8). Fumarase catalyzes the conversion of fumarate to malate (9). Fumarase deficiency leads to the accumulation of fumarate, an oncometabolite that has been shown to promote epithelial-to-mesenchymal-transition (EMT), a developmental process that has been implicated in oncogenesis (10).

This kit contains reagents to examine total protein levels of the five NF-κB/Rel family members: p65/RelA, RelB, c-Rel, NF-κB1 (p105/p50) and NF-κB2 (p100/p52).

Background: Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, c-Rel, RelB, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Both p105 and p100 are proteolytically processed by the proteasome to produce p50 and p52, respectively. Rel proteins bind p50 and p52 to form dimeric complexes that bind DNA and regulate transcription. In unstimulated cells, NF-κB is sequestered in the cytoplasm by IκB inhibitory proteins (3-5). NF-κB-activating agents can induce the phosphorylation of IκB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-κB to enter the nucleus where it regulates gene expression (6-8). NIK and IKKα (IKK1) regulate the phosphorylation and processing of NF-κB2 (p100) to produce p52, which translocates to the nucleus (9-11).

The IRAK Isoform Antibody Sampler Kit provides an economical means to examine total protein levels of the four Interleukin-1 Receptor Associated Kinase family members: IRAK1, IRAK2, IRAK3/IRAK-M, and IRAK4.

Background: Interleukin-1 (IL-1) receptor-associated kinase (IRAK) is a serine/threonine-specific kinase that can be coprecipitated in an IL-1-inducible manner with the IL-1 receptor (1). The mammalian family of IRAK molecules contains four members (IRAK1, IRAK2, IRAK3/IRAK-M, and IRAK4). The binding of IL-1 to IL-1 receptor type I (IL-1RI) initiates the formation of a complex that includes IL-1RI, AcP, MyD88, and IRAKs (2). IRAK undergoes autophosphorylation shortly after IL-1 stimulation. The subsequent events involve IRAK dissociation from the IL-1RI complex, its ubiquitination, and its association with two membrane-bound proteins: TAB2 and TRAF6. The resulting IRAK-TRAF6-TAB2 complex is then released into the cytoplasm where it activates protein kinase cascades, including TAK1, IKKs, and the stress-activated kinases (3).

The Class I HDAC Antibody Sampler Kit provides an economical means of detecting Class I HDAC proteins using control antibodies against HDAC1, HDAC2 and HDAC3. The kit contains enough primary antibodies to perform at least two western blot experiments.

Background: Acetylation of the histone tail causes chromatin to adopt an "open" conformation, allowing increased accessibility of transcription factors to DNA. The identification of histone acetyltransferases (HATs) and their large multiprotein complexes has yielded important insights into how these enzymes regulate transcription (1,2). HAT complexes interact with sequence-specific activator proteins to target specific genes. In addition to histones, HATs can acetylate nonhistone proteins, suggesting multiple roles for these enzymes (3). In contrast, histone deacetylation promotes a "closed" chromatin conformation and typically leads to repression of gene activity (4). Mammalian histone deacetylases can be divided into three classes on the basis of their similarity to various yeast deacetylases (5). Class I proteins (HDACs 1, 2, 3, and 8) are related to the yeast Rpd3-like proteins, those in class II (HDACs 4, 5, 6, 7, 9, and 10) are related to yeast Hda1-like proteins, and class III proteins are related to the yeast protein Sir2. Inhibitors of HDAC activity are now being explored as potential therapeutic cancer agents (6,7).

The NF-κB Family Antibody Sampler Kit II provides an economical means of detecting members of the NF-κB family. The kit includes enough antibody to perform two western blots with each primary antibody.

Background: Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, c-Rel, RelB, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Both p105 and p100 are proteolytically processed by the proteasome to produce p50 and p52, respectively. Rel proteins bind p50 and p52 to form dimeric complexes that bind DNA and regulate transcription. In unstimulated cells, NF-κB is sequestered in the cytoplasm by IκB inhibitory proteins (3-5). NF-κB-activating agents can induce the phosphorylation of IκB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-κB to enter the nucleus where it regulates gene expression (6-8). NIK and IKKα (IKK1) regulate the phosphorylation and processing of NF-κB2 (p100) to produce p52, which translocates to the nucleus (9-11).

The Histone Deacetylase (HDAC) Antibody Sampler Kit provides a fast and economical means to evaluate the endogenous levels of HDACs. The kit contains enough primary and secondary antibodies to perform two Western blot experiments.

Background: Acetylation of the histone tail causes chromatin to adopt an "open" conformation, allowing increased accessibility of transcription factors to DNA. The identification of histone acetyltransferases (HATs) and their large multiprotein complexes has yielded important insights into how these enzymes regulate transcription (1,2). HAT complexes interact with sequence-specific activator proteins to target specific genes. In addition to histones, HATs can acetylate nonhistone proteins, suggesting multiple roles for these enzymes (3). In contrast, histone deacetylation promotes a "closed" chromatin conformation and typically leads to repression of gene activity (4). Mammalian histone deacetylases can be divided into three classes on the basis of their similarity to various yeast deacetylases (5). Class I proteins (HDACs 1, 2, 3, and 8) are related to the yeast Rpd3-like proteins, those in class II (HDACs 4, 5, 6, 7, 9, and 10) are related to yeast Hda1-like proteins, and class III proteins are related to the yeast protein Sir2. Inhibitors of HDAC activity are now being explored as potential therapeutic cancer agents (6,7).

The Parkinson's Research Antibody Sampler Kit provides an economical means of detecting target proteins related to Parkinson's disease. The kit contains enough primary and secondary antibody to perform two western blots per primary.

Background: Autophagy is a catabolic process for the autophagosome-lysosomal degradation of bulk cytoplasmic contents (1, 2). Selective autophagy targets the degradation of distinct sets of substrates and organelles (3-5). One of the best studied examples of selective autophagy involves the clearance of damaged mitochondria through a process called mitophagy. Several pathways have been described for various contexts of mitophagy, including the FUNDC1 pathway, the BNIP3 and BNIP3L/Nix pathway, and the PINK1/Parkin pathway. FUNDC1 is a mitochondrial protein that is phosphorylated by the autophagy kinase ULK1 and regulates hypoxia induced mitophagy (6, 7). BNIP3L/Nix and BNIP3 are members of the Bcl-2 family of apoptosis regulators that are expressed on mitochondria, induced by hypoxia, and have have been shown to play a role in mitophagy (8). BNIP3L/Nix is also important in the autophagic maturation of erythroid cells (9). FUNDC1, BNIP3 and BNIP3L/Nix bind to LC3 family members, targeting the mitochondria to the autophagosome.Non-hypoxic induction of mitophagy can be regulated by the PINK1/Parkin pathway, which plays causative roles in neurodegenerative disease, most notably Parkinson’s disease (10, 11). PINK1 is a mitochondrial serine/threonine kinase that is stabilized on the outer mitochondrial membrane of damaged mitochondria. Substrates of PINK1 include the E3 ubiquitin ligase Parkin and ubiquitin itself (12-14). Phosphorylation of Parkin as well as binding to phosphorylated ubiquitin leads to accumulation of ubiquitinated chains on multiple mitochondrial proteins. Ubiquitinated proteins are recognized by selective cargo receptors including SQSTM1/p62, Optineurin, and NDP52 (15-16). Autophagy cargo receptors contain an LC3-interacting region (LIR) required for binding to Atg8/LC3 family members and targeting to the autophagosome (3).